Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach
Protein optimization can generally be obtained by rational enzyme design and directed evolution. Phage display has been used as a method for directed evolution. Trypsin has been studied as a biocatalyst in peptide synthesis despite its usual function in hydrolyzing peptide bonds. The objective of th...
| Main Authors: | , , |
|---|---|
| Format: | Proceeding Paper |
| Language: | English English |
| Published: |
2015
|
| Subjects: | |
| Online Access: | http://irep.iium.edu.my/48397/ http://irep.iium.edu.my/48397/1/Badri%20KOP.pdf http://irep.iium.edu.my/48397/2/Poster_badri_ACB2015.pdf |
| _version_ | 1848783292844736512 |
|---|---|
| author | Abdul Kudos, Muhammad Badri Gissel, Sabrina Bordusa, Frank |
| author_facet | Abdul Kudos, Muhammad Badri Gissel, Sabrina Bordusa, Frank |
| author_sort | Abdul Kudos, Muhammad Badri |
| building | IIUM Repository |
| collection | Online Access |
| description | Protein optimization can generally be obtained by rational enzyme design and directed evolution. Phage display has been used as a method for directed evolution. Trypsin has been studied as a biocatalyst in peptide synthesis despite its usual function in hydrolyzing peptide bonds. The objective of this study is to identify trypsin variants with a more hydrophobic cluster around its active site, potentially yielding higher peptide synthesis products rather than its hydrolysis competitive counterpart. A trypsin variant library displayed by phages with randomization in 5 amino acids located around the active site of the trypsin variant K60E/N143H/E151H/D189K is selected based on a designed immobilization-elution-assay in regard the transamidation reaction model. A preselection stage of recombinant phages was done with MyCUT tag, binding to the immobilized anti-c-myc antibodies. 2 trypsin variants were successfully expressed, purified, characterized and sequenced. Both of them gave a higher accumulative hydrophobicity relative index. This study has shown that selection of a potentially optimized trypsin variant with a more hydrophobic cluster at active site to reduce the hydrolysis products is possible by using the phage display approach. |
| first_indexed | 2025-11-14T16:19:02Z |
| format | Proceeding Paper |
| id | iium-48397 |
| institution | International Islamic University Malaysia |
| institution_category | Local University |
| language | English English |
| last_indexed | 2025-11-14T16:19:02Z |
| publishDate | 2015 |
| recordtype | eprints |
| repository_type | Digital Repository |
| spelling | iium-483972017-06-01T08:12:33Z http://irep.iium.edu.my/48397/ Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach Abdul Kudos, Muhammad Badri Gissel, Sabrina Bordusa, Frank TP248.13 Biotechnology Protein optimization can generally be obtained by rational enzyme design and directed evolution. Phage display has been used as a method for directed evolution. Trypsin has been studied as a biocatalyst in peptide synthesis despite its usual function in hydrolyzing peptide bonds. The objective of this study is to identify trypsin variants with a more hydrophobic cluster around its active site, potentially yielding higher peptide synthesis products rather than its hydrolysis competitive counterpart. A trypsin variant library displayed by phages with randomization in 5 amino acids located around the active site of the trypsin variant K60E/N143H/E151H/D189K is selected based on a designed immobilization-elution-assay in regard the transamidation reaction model. A preselection stage of recombinant phages was done with MyCUT tag, binding to the immobilized anti-c-myc antibodies. 2 trypsin variants were successfully expressed, purified, characterized and sequenced. Both of them gave a higher accumulative hydrophobicity relative index. This study has shown that selection of a potentially optimized trypsin variant with a more hydrophobic cluster at active site to reduce the hydrolysis products is possible by using the phage display approach. 2015 Proceeding Paper NonPeerReviewed application/pdf en http://irep.iium.edu.my/48397/1/Badri%20KOP.pdf application/pdf en http://irep.iium.edu.my/48397/2/Poster_badri_ACB2015.pdf Abdul Kudos, Muhammad Badri and Gissel, Sabrina and Bordusa, Frank (2015) Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach. In: ASIAN Congress on Biotechnology 2015, 15-19 Nov 2015, Kuala Lumpur. (Unpublished) |
| spellingShingle | TP248.13 Biotechnology Abdul Kudos, Muhammad Badri Gissel, Sabrina Bordusa, Frank Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach |
| title | Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach |
| title_full | Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach |
| title_fullStr | Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach |
| title_full_unstemmed | Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach |
| title_short | Selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach |
| title_sort | selection of potentially optimized trypsin variant with a more hydrophobic cluster active site via a phage display approach |
| topic | TP248.13 Biotechnology |
| url | http://irep.iium.edu.my/48397/ http://irep.iium.edu.my/48397/1/Badri%20KOP.pdf http://irep.iium.edu.my/48397/2/Poster_badri_ACB2015.pdf |